I. Field
The present disclosure pertains to methods and compositions for producing single chain multivalent binding proteins that specifically bind to one or more desired target antigens. More specifically, the disclosure relates to protein, nucleic acid, and cellular libraries of single chain multivalent binding proteins (e.g., scDVD molecules) and methods of using these libraries for the screening of single chain multivalent binding proteins using cell surface display technology (e.g., yeast display).
II. Description of Related Art
A wide variety of multispecific antibody formats have been developed (see Kriangkum, J., et al., Biomol Eng, 2001. 18(2): p. 31-40). Amongst them tandem single-chain Fv molecules and diabodies, and various derivatives there of, are the most widely used formats for the construction of recombinant bispecific antibodies. More recently diabodies have been fused to Fc to generate more Ig-like molecules, named di-diabodies (see Lu, D., et al., J Biol Chem, 2004. 279(4): p. 2856-65). In addition, multivalent antibody construct comprising two Fab repeats in the heavy chain of an IgG and capable of binding four antigen molecules has been described (see WO 0177342A1, and Miller, K., et al., J Immunol, 2003. 170(9): p. 4854-61).
Despite the many bispecific antibody formats available to the skilled artisan, there is often a need for the skilled artisan to improve the affinity of the bispecific antibody through affinity maturation. However, conventional affinity maturation approaches rely upon screening for affinity matured variants of the component binding domains of the multispecific antibody followed by their reassembly into the original multispecific format. Such reassembly often results in a loss of the desired improvement in binding affinity or other desirable binding characteristics. Accordingly, there is a need in the art for improved constructs, formats, and screening methodologies for identifying affinity variants of multivalent binding proteins in their desired multivalent format.